Since NFT are the most striking pathological feature in tauopathies, much attention has focused on understanding how the deposition of NFT may cause neurodegeneration, in essence using animal models recapitulating NFT pathology to investigate the mechanism of disease. It has long been postulated that the aggregation of tau into filaments and NFT results in a toxic gain of function. This view has been substantially challenged by observations that neuronal loss and memory impairment can experimentally be cured despite ongoing NFT formation (Santacruz et al. 2005). In some animal models, the tau-mediated loss of neurons does not even require NFT development. Thus, it is assumed that non-filamentous but already aggregated, globular tau intermediates on route to assemble into larger helical filaments may represent the neurotoxic tau species.
Elevated levels of free tau, not bound to microtubules, presumably increases its likelihood to become misfolded, as well as undergo modifications or conformational changes that promote the formation of aggregated small globular oligomers that eventually will assemble into insoluble filaments. Covalent modifications stabilizing conformational changes most likely include phosphorylation since tau protein isolated from AD brain was found to be abnormally high phosphorylated at multiple critical sites (‘hyperphosphorylated tau’) and it was demonstrated that pseudo-hyperphosphorylation (i.e. pseudo-phosphorylated at multiple sites along the protein) can facilitate abnormal conformation of tau protein (Jeganathan et al. 2008). In this context tau hyperphosphorylation was identified as diagnostic target (WO9311231A1) and different tau related Kinases identified as therapeutic target (WO2007088400A1).
Given that tau is normally a highly soluble protein that does not readily aggregate, this matter has been difficult to assess in experimental models because of the resistance of tau to aggregate within an ideal time-frame for culture studies or within an animal's relatively short lifespan. Because high concentrations of tau are required to promote tau aggregation in experimental models, it is believed that the enhanced ability of tau to form small globular aggregates in the cytoplasm of neurons and glia in human tauopathies may be due to pathological conditions that locally increase the pool of tau available for aggregation. Yet it is unlikely that the amount of tau in various tauopathies is as high as in cell culture and animal models that artificially force massive tau overexpression and, therefore, much caution is needed extrapolating results from such model systems to the human condition.
Further complicating matters is evidence that mouse tau appears to prevent tau aggregation in transgenic mice overexpressing wild-type human tau (htau). Nonetheless, transgenic mice that overexpress high levels of htau isoforms containing aggregation-promoting mutations (e.g. P301L tau) can develop tau pathology even in the presence of endogenous mouse tau. The P301L and P301S mutations are among the first described FTD mutations and show a very early mean onset for FTD in man. Tau transgenic mouse models with expression of these mutants display onset of first signs of tau pathology starting at 2.5 to 5 months (Schindowski et al. 2006). WO 01/53340 A2 discloses mouse models expressing wildtype or tau with one mutation like the named P301L mutation for generating a neurodegenerative disease model as well as tool for drug development. Furthermore a transgenic mouse model with tau cDNA bearing three FTD mutations the tau pathology has been described (Lim et al., 2001).
To accelerate tau aggregation in vitro, polyanionic cofactors or small molecule ligands are often used to facilitate tau aggregation. For example, in a cell culture model overexpressing full length tau, Congo red treatment stimulates the formation of filamentous tau aggregates and decreases cell viability (Bandyopadhyay et al. 2007). These and other results suggest that also in cell culture models tau aggregation causes cell death or, at least, accelerates its onset. However, no cell culture model has been described so far that does not force aggregation of tau by either artificial high concentrations of tau or addition of, at higher doses toxic, compounds in order to facilitate or precipitate aggregation (Ko et al. 2004, Tsukane et al. 2007, Nie et al. 2007). In view of modelling tauopathy disease mechanisms for drug development purposes, both strategies bear a high risk of producing artificial results since the mechanism of degeneration may significantly differ from tau pathology in the AD brain.